ABSTRACT
Wastewater surveillance using RT-qPCR has now been widely adopted to track circulating levels of SARS-CoV-2 virus in many sewer sheds. The CDC qPCR assays targeting two regions (N1 and N2) within the N gene are commonly used, but a discrepancy between the two biomarkers has been noticed by many groups using this method since late 2021. The reason is presumed to be due to mutations in regions targeted by the qPCR probe. In this study, we systematically investigated and unequivocally confirmed that the underlying reason for this discrepancy was mutations in the N1 probe target, and that a single mutation could cause a significant drop in signal. We first confirmed the proportion of related mutations in wastewater samples (Jan 2021-Dec 2022) using nested PCR and LC-MS. Based on relative proportion of N1 alleles, we separated the wastewater data into four time periods corresponding to different variant waves: Period I (Alpha and Delta waves with 0 mutation), Period II (BA.1/BA.2 wave with a single mutation found in all Omicron strains), Period III (BA. 5.2* wave with two mutations), and Period IV (BQ.1* wave with two mutations). Significantly lower N1 copies relative to N2 copies in samples from Periods II-IV compared to those from Period I was observed in wastewater. To further pinpoint the extent to which each mutation impacted N1 quantification, we compared the qPCR response among different synthetic oligomers with corresponding mutations. This study highlighted the impact of even just one or two mutations on qPCR-based wastewater surveillance of SARS-CoV-2.
ABSTRACT
Background: While the emergence of the Coronavirus Disease 2019 (COVID-19) has greatly affected the mental health and wellbeing of people everywhere, the core symptoms of common syndromes of psychological distress remain unclear, especially among COVID-19 patients who have been quarantined in Square Cabin Hospitals. Using the method of network analysis, the current study explored the relations between stress, depression, anxiety and insomnia symptoms among COVID-19 patients in Shanghai Four-Leaf Clover Cabin Hospital. Method: The study used an online survey to gauge the mental health issues of COVID-19 patients at Shanghai Four-Leaf Clover Cabin Hospital from April 18 to May 19, 2022. The Perceived Stress Scale, Patient Health Questionnaire, Generalized Anxiety Disorder Scale, and Insomnia Severity Index were used to assess perceived stress symptoms, depressive symptoms, anxiety symptoms, and insomnia symptoms, respectively. The central symptoms and bridge symptoms were determined using a regularized partial correlation network. Results: A total of 1014 patients was included in this study. Overall, 61.5%, 49.2% , and 56.1% of all patients reported symptoms of depressive, anxiety, and insomnia, respectively. “Distress caused by the sleep difficulties” had the highest expected influence, followed by “Excessive worry”, “Severity of sleep onset”, “Overburden”, “Uncontrollable worry”, “Trouble relaxing”, “Restlessness”. Seven bridge symptoms were identified: “Sad Mood”, “Severity of sleep onset”, “Sleep”, “Motor”, “Feeling afraid”, “Nervousness”, “Appetite”. In addition, significant differences in network global strength were found between women and men. Conclusions: Psychological distress was prevalent among COVID-19 patients and insomnia symptoms played a key role in the network of psychological distress symptoms. As a result, specific insomnia symptoms should be closely monitored as potential intervention targets in alleviating or preventing common, frequently co-occurring psychiatric syndromes.